Relations between the methods of isolating Brucella lipopolysaccharides and their effect on hemopoiesis in mice

The comparative study of the effect produced by different lipopolysaccharide (LPS) preparations obtained from B. melitensis virulent strain 565 and B. abortus vaccine strain 19-BA on hematopoiesis in mice was made. The LPS preparations were obtained (1) by Boivin's technique, (2) by Westphal's technique and (3) by mild alkaline hydrolysis of Bovin's active complex, this technique having been developed at the Brucellosis Laboratory of the Gamaleya Research Institute of Epidemiology and Microbiology. All tests (the spleen endocolonization test, the hydroxyurea kill test, the determination of the content of splenic colony-forming units in the peripheral blood) showed that LPS from B. melitensis virulent strain 565 had a more pronounced disturbing effect on hematopoiesis than LPS from B. abortus vaccine strain 19-BA. Among the LPS preparations obtained by different methods, the one obtained with the use of the technique developed at the Gamaleya Research Institute of Epidemiology and Microbiology proved to have the mildest effect on hematopoiesis, probably due to the partial saponification of the lipid component of LPS. Lipid A in a dose of 0.1-10 micrograms produced no activating effect on the hematopoiesis characteristics under study. None of the LPS preparations proved to be capable of stimulating the formation of transitory endogenous colonies in the spleen of mice.     

An overview of malaria transmission from the perspective of Amazon Anopheles vectors

In the Americas, areas with a high risk of malaria transmission are mainly located in the Amazon Forest, which extends across nine countries. One keystone step to understanding the Plasmodium life cycle in Anopheles species from the Amazon Region is to obtain experimentally infected mosquito vectors. Several attempts to colonise Ano- pheles species have been conducted, but with only short-lived success or no success at all. In this review, we review the literature on malaria transmission from the perspective of its Amazon vectors. Currently, it is possible to develop experimental Plasmodium vivax infection of the colonised and field-captured vectors in laboratories located close to Amazonian endemic areas. We are also reviewing studies related to the immune response to P. vivax infection of Anopheles aquasalis, a coastal mosquito species. Finally, we discuss the importance of the modulation of Plasmodium infection by the vector microbiota and also consider the anopheline genomes. The establishment of experimental mosquito infections with Plasmodium falciparum, Plasmodium yoelii and Plasmodium berghei parasites that could provide interesting models for studying malaria in the Amazonian scenario is important. Understanding the molecular mechanisms involved in the development of the parasites in New World vectors is crucial in order to better determine the interaction process and vectorial competence.     

Genetic diversity of chloroquine-resistant Plasmodium vivax parasites from the western Brazilian Amazon

The molecular basis of Plasmodium vivax chloroquine (CQ) resistance is still unknown. Elucidating the molecular background of parasites that are sensitive or resistant to CQ will help to identify and monitor the spread of resistance. By genotyping a panel of molecular markers, we demonstrate a similar genetic variability between in vitro CQ-resistant and sensitive phenotypes of P. vivax parasites. However, our studies identified two loci (MS8 and MSP1-B10) that could be used to discriminate between both CQ-susceptible phenotypes among P. vivax isolates in vitro. These preliminary data suggest that microsatellites may be used to identify and to monitor the spread of P. vivax-resistance around the world.     

Dissociative mechanism of thermal denaturation of rabbit skeletal muscle glycogen phosphorylase b

The thermal stability of rabbit skeletal muscle glycogen phosphorylase b was characterized using enzymological inactivation studies, differential scanning calorimetry, and analytical ultracentrifugation. The results suggest that denaturation proceeds by the dissociative mechanism, i.e., it includes the step of reversible dissociation of the active dimer into inactive monomers and the following step of irreversible denaturation of the monomer. It was shown that glucose 1-phosphate (substrate), glucose (competitive inhibitor), AMP (allosteric activator), FMN, and glucose 6-phosphate (allosteric inhibitors) had a protective effect. Calorimetric study demonstrates that the cofactor of glycogen phosphorylase-pyridoxal 5'-phosphate-stabilizes the enzyme molecule. Partial reactivation of glycogen phosphorylase b preheated at 53 degrees C occurs after cooling of the enzyme solution to 30 degrees C. The fact that the rate of reactivation decreases with dilution of the enzyme solution indicates association of inactive monomers into active dimers during renaturation. The allosteric inhibitor FMN enhances the rate of phosphorylase b reactivation.     

Comparative cytogenetics of hamsters of the genus Calomyscus

Karyotypes of Calomyscus from different regions of Turkmenistan, Iran, and Azerbaijan were studied using chromosome banding (G- and C-banding) and analyses of meiosis in laboratory hybrids. Extensive variation in the diploid number and the number of autosomal arms (FNa) was revealed (2n = 30, FNa = 44; 2n = 32, FNa = 42; 2n = 44, FNa = 46; 2n = 44, FNa = 58; 2n = 37, FNa = 44; 2n = 50, FNa = 50; 2n = 52, FNa = 56). Centric and tandem fusions and heterochromatin changes were identified as the major modes of karyotype evolution in this group. Natural hybrids between individuals with different karyotypes were recorded, and regular chromosome pairing in meiosis was observed in laboratory hybrids. Fluorescence in situ hybridization with a 353-bp BspRI complex tandem repeat indicated that chromosomal repatterning occurred recently within the genus. There is no unequivocal evidence suggesting the role of chromosomal change in the speciation of the populations of Calomyscus examined.     

Genetic diversity of Chionomys genus (Mammalia, Arvicolinae) and comparative phylogeography of snow voles

In the present study, the genetic polymorphism of the Chionomys genus was examined based on the sequencing of the mitochondrial cytb gene and two nuclear exons, including GHR exon 10 and BRCA1 exon 11. The distinct subdivision of the genus of snow voles into five lineages, including Ch. nivalis, Ch. gud, Ch. roberti, and Ch. aff. nivalis from Turkey, as well as Ch. aff. gud from Turkey, was demonstrated. The branching order in the trees constructed based on the data for different genes was ambiguous, which was probably the consequence of recent and rapid radiation of the major lineages from a common ancestor. However, the data of the mitochondrial and nuclear gene analyses definitely indicated that the genetic and taxonomic diversity of the Chionomys genus was higher than it was expected before. The genetic divergence of some populations was so deep that they probably deserved the statuses of independent species. Despite that the range of the European snow vole Ch. nivalis is larger and more fragmented than the Gudaur vole Ch. gud, the latter species with its relatively small range, which is limited to the Caucasian and Pontic Mountains, was characterized by a similarly expressed phylogenetic structure. At the same time, Robert’s vole Ch. roberti was less structured genetically than the first two species. The data obtained supported the Near Eastern, rather than the European origin of the Chionomys genus.     

In vitro and in vivo conditional sensitization of hepatocellular carcinoma cells to TNF-induced apoptosis by Taxol

High mortality among hepatocellular carcinoma (HCC) patients reflects both late diagnosis and low curability, due to pharmacoresistance. Taxol (TAX) is toxic for many human HCC-derived cell lines, yet its clinical efficacy on HCCs is poor. Combining TAX with other drugs appears a promising possibility to overcome such refractoriness. We analyzed whether combining tumor necrosis factor (TNF) with TAX would improve their toxicity. Human HCC-derived cell lines were treated with TAX or TNF, alone or combined. Apoptosis was assessed by morphology and flow-cytometry. Several pro- and anti-apoptotic molecules were evaluated by western blotting and/or enzymatic assay. After a 24 hour treatment, TNF was ineffective and TAX modestly cytotoxic, whereas HCC cells were conditionally sensitized to TNF by TAX. Indeed some relevant parameters were shifted to a prodeath setting: TNF-receptor 1 was increased, SOCS3, c-FLIP and pSTAT3 were markedly downregulated. These observations provide a significant clue to critically improve the drug susceptibility of HCC cells by combining 2 agents, TAX and TNF. The sequential application of TAX at a low dosage followed by TNF for only a short time triggered a strong apoptotic response. Of interest, prior TAX administration could also sensitize to TNF-induced apoptosis in the Yoshida AH-130 hepatoma transplanted in mice. Therefore, scrutinizing the possibility to develop similar combination drug regimens in suitable preclinical models seems highly advisable.